GSM7473748: HK2-Hypoxia_Input; Homo sapiens; ChIP-Seq
Cells were fixed with 1% formaldehyde at room temperature for 10 min and quenched in 2.5 M glycine (1:20) for 5 min. Cell precipitates were collected and at centrifuged with 1200 r/min and at 4 °C for 3min. The collected cell precipitates were then sonicated to generate DNA fragments. Chromatin fragments were immunoprecipitated and incubated at 4 °C overnight using antibodies against HIF-1α (ab243860, Abcam). 50 μl magnetic beads (Protein G) were placed into 1.5ml EP tubes and mixed with chip binding buffer, and the supernatant was not discarded until liquid was transparent. Then, chromatin fragments-antibody mixture was added with magnetic beads and incubated for 4 h at 4 °C. The precipitated DNA was purified using the ChIP DNA Clean Kit. Chip sequencing was conducted by KeyGEN Bio TECH. Sequencing was performed using VAHTS mRNA-seq V3 Library Prep Kit for Illumina NovaSeq 6000.